We have recently observed that the third component of human complement (C3) becomes fixed to the red blood cell (RBC) membrane during storage of blood at 4 degrees C. Specifically, the C3 on the stored RBCs contains the C3c fragment. We propose to extend these observations and study the role of membrane-bound C3 fragment in the removal mechanism of the nonviable, stored RBCs following transfusion. We also propose to study platelet-comlement interaction during 4 degrees C and 22 degrees C storage to explore the role of complement in the removal of stored platelets in-vivo. These studies will involve quantitation of RBC and platelet membrane-bound C3 fragment utilizing a new developed method. This new method should improve the accuracy of the quantitation of the cell-bound C3 over the previously published methods. Our method is based on the ability of bound C3 to neutralize a proportion of a "standard" amount of anti-C3c antibody. A set of standard RBCs having known amounts of C3 will be prepared, and the anti-C3c antibody will be assayed by an automated hemagglutination method. The new quantitative assay will be used also to study RBCs of patients with certain hemolytic anemias. These studies will involve performance of in vitro phagocytosis assay and in-vivo RBC survival studies. The latter studies are designed to evaluate the correlation between the percentage viable RBCs in stored blood and the quantity of C3 fragment bound to the RBC membrane. Since it is known that ATP concentration correlates with RBC viability, we will transfuse stored, ATP-regenerated RBCs. We will also use differential agglutination technics to recover physically the previously stored RBCs after transfusion and measure the rate of change of ATP and of the quantity of C3 fragment in these recovered RBCs to evaluate the association of these compounds with the removal of nonviable RBCs. In addition, we will attempt to clarify the mechanism of C3 attachment to RBCs during storage and to identify C3 fragment involved.